Lactobacilli have previously been used to deliver vaccine components for active immunization in vivo. Vectors encoding a single-chain Fv (scFv) antibody fragment, which recognizes the streptococcal antigen I/II (SAI/II) adhesion molecule of Streptococcus mutans, were constructed and expressed in Lactobacillus zeae (American Type Culture Collection (ATCC) 393). The scFv antibody fragments secreted into the supernatant or expressed on the surface of the bacteria showed binding activity against SAI/II in enzyme-linked immunosorbent assay (ELISA), and surface scFv-expressing lactobacilli agglutinated SAI/II-expressing S. mutans in vitro without affecting the corresponding SAI/II knockout strain. Lactobacilli expressing the scFv fragment fused to an E-tag were visualized by scanning electron microscopy (SEM) using beads coated with a monoclonal anti-E-tag antibody, and they bound directly to beads coated with SAI/II. After administration of scFv-expressing bacteria to a rat model of dental caries development, S. mutans bacteria counts and caries scores were markedly reduced. As lactobacilli are generally regarded as safe (GRAS) microorganisms, this approach may be of considerable commercial interest for in vivo immunotherapy.

Background:The role of Hsf1 in mammary tumorigenesis and metastasis remains elusive. Results: Hsf1 deletion inhibits mammary tumorigenesis and metastasis by reducing ERK1/2 activity and epithelial-mesenchymal transition of mammary epithelial cells. Conclusion: Deletion of Hsf1 in mice carrying Her2/Neu significantly reduces breast cancer and metastasis. Significance: These findings indicate a powerful inhibitory effect conferred by Hsf1 in ErbB2-induced breast cancer.

Heat shock factor 1 (HSF1) regulates the rapid and transient expression of heat shock genes in response to stress. The transcriptional activity of HSF1 is tightly controlled, and under physiological growth conditions, the HSF1 monomer is in a heterocomplex with the molecular chaperone HSP90. Through unknown mechanisms, transcriptionally repressed HSF1⅐HSP90 heterocomplexes dissociate following stress, which triggers HSF1 activation and heat shock gene transcription. Using a yeast two-hybrid screening system, we have identified Ral-binding protein 1 (RalBP1) as an additional HSF1-interacting protein. We show that RalBP1 and HSF1 interact in vivo, and transient cotransfection of HSF1 and RalBP1 into hsf1 ؊/؊ mouse embryo fibroblasts represses HSP70 expression. Furthermore, transient cotransfection of HSF1 and the constitutively active form of RalA (RalA23V), an upstream activator of the RalBP1 signaling pathway, increases the heatinducible expression of HSP70, whereas the dominant negative form (RalA28N) suppresses HSP70 expression. We further find that ␣-tubulin and HSP90 are also present in the RalBP1⅐HSF1 heterocomplexes in unstressed cells. Upon heat shock, the Ral signaling pathway is activated, and the resulting RalGTP binds RalBP1. Concurrently, HSF1 is activated, leaves the RalBP1⅐HSF1⅐HSP90⅐␣-tubulin heterocomplexes, and translocates into the nucleus, where it then activates transcription. In conclusion, these observations reveal that the RalGTP signal transduction pathway is critical for activation of the stress-responsive HSF1 and perhaps HSP90 molecular chaperone system.Mammalian heat shock factor 1 (HSF1), 1 a phosphorylated protein, regulates the stress inducibility of heat shock genes. Phosphorylation of HSF1 is indicative of its complex mode of regulation by various signaling pathways. Studies using phosphopeptide analysis of HSF1 protein as well as studies analyzing the transactivation properties of HSF1 using chimeric constructs containing GAL4-HSF1 or LexA-HSF1 have suggested that phosphorylation of serine residues Ser 303 , Ser 307 , and Ser 363 is likely to be involved in repression of HSF1 transcriptional activity (1-7). Mitogen-activated protein kinases and glycogen synthase kinase 3 are candidates for phosphorylating these residues. HSF1 could potentially be phosphorylated during its activation process as well, perhaps at Ser 230 by calcium calmodulin protein kinase II (8). HSF1 is also found in multichaperone complexes under physiological conditions and during its repression (9 -11). Specifically, HSP90 has been coimmunoprecipitated with the monomeric form of HSF1, suggesting that an HSP90⅐HSF1 heterocomplex may keep HSF1 in a repressed state. Disruption of this heterocomplex by stress would allow HSF1 to form trimers and acquire DNA binding capability. One likely outcome of the disruption of HSF1⅐HSP90 heterocomplexes during stress is the accumulation of denatured polypeptides and the ability of HSP90 to bind such denatured polypeptides (9 -11). During recovery from stress, HSF1 trimers h...

Acute kidney injury (AKI) is associated with mitochondrial fragmentation, which contributes to mitochondrial damage and tubular cell apoptosis. Mitochondrial fragmentation involves the cleavage of both mitochondrial outer and inner membranes. Cleavage of the outer membrane results from Drp-1-mediated fission activation and Bak-promoted fusion arrest, but the molecular mechanism of inner membrane cleavage remains elusive. OMA1-mediated proteolysis of OPA1, a key inner membrane fusion protein, was recently suggested to account for inner membrane cleavage during cell stress. In this study, we determined the role of OMA1 in OPA1 proteolysis and mitochondrial fragmentation in experimental models of ischemic AKI. In ATP-depletion injury, knockdown of OMA1 suppressed OPA1 proteolysis, mitochondrial fragmentation, cytochrome c release, and consequent apoptosis in renal proximal tubular cells. In mice, OMA1 deficiency prevented ischemic AKI as indicated by better renal function, less tubular damage, and lower apoptosis. OPA1 proteolysis and mitochondrial injury during ischemic AKI were ameliorated in OMA1-deficient mice. Thus, OMA1-mediated OPA1 proteolysis plays an important role in the disruption of mitochondrial dynamics in ischemic AKI.

The heat shock transcription factors (Hsfs) activate the stress-inducible expression of heat shock proteins (Hsps) and other molecular chaperones in response to stress and, therefore, play an essential role in protein disaggregation and protein folding. In humans, missense mutation in the hsf4 gene causes cataract, and mice bearing a targeted disruption of the hsf4 gene exhibit defects in lens fiber cell differentiation and early cataract formation. Here, we show that Hsf4b is a direct target of the mitogen-activated protein (MAP) kinase extracellular signal-related kinase (ERK) and that phosphorylation of Hsf4b by ERK leads to increased ability of Hsf4b to bind DNA. Surprisingly, Hsf4b also interacts with an ERK-specific dual-specificity tyrosine phosphatase named DUSP26 identified from a yeast two-hybrid screen. While activated ERK phosphorylates Hsf4b, DUSP26 controls the activity of ERK, leading to phosphorylation/dephosphorylation of Hsf4b, altering its ability to bind DNA. Therefore, DUSP26 interaction with Hsf4b places this transcription factor within a regulatory circuit in the MAP kinase signaling pathway.